The purpose of the research described in this proposal is to identify and characterize protective immune responses against defined and newly discovered Plasmodium vivax antigens from the pre-erythrocytic and mosquito stages of the parasite., and then to formulate vaccines that induce these immune responses and provide protection in the Aotus monkey model. (A) Immune responses thought to be involved in protection will be identified in residents of endemic areas with life-long exposure to P. vivax malaria and also in volunteers immunized with irradiated P. vivax sporozoites and shown to be protected against sporozoite challenge. (b) Testing for protective biological activity will be done in vivo by assessing parasitemia in Aotus monkeys and in human volunteers, and in vitro by assessing the ability of sera to inhibit sporozoite invasion of hepatocytes, development of sporozoites in hepatocyte culture, and development of blood stages in blood stage culture, and by assessing oocyst development in Anopheles albimanus fed on blood stage cultures containing gametocytes. (c) Sera from protected and non protected individuals will be characterized by sporozoite, liver and blood stage IFAT and by ELISA utilizing selected full length recombinant proteins, long peptides or components epitopes. We will focus initially on reagents that are available: PvSP and PvSSP2/TRAP (sporozoite and liver stage antigens), PvMSP-1 and PvDBP (blood stage antigens) and Pvs25/28 (antigens expressed in the mosquito gut). Other antigens will be studied as they become available. Lymphocytes will be characterized by CTL and interferon gamma production by ELISPOT in response to epitopes derived from the PvCSP and PvSSP2 (and from proteins or peptides formulated with various adjuvants, DNA plasmids, and recombinant poxviruses, and monkey models. We will utilize the blood stage challenge model in Aotus and a newly developed sporozoite challenge model in Aotus. This research project will integrate the use of laboratories, primate facilities, insectories and field sites painstakingly developed over the last decade in western Columbia in order to answer these critical biological questions about this important but relatively neglected malaria parasite.